Some viral vectors have been shown to effectively induce immunity to a transgene product. This property is being exploited to develop novel vaccines based on gene transfer of autologous antigens, such as tumor-associated antigens, or exogenous antigens, such as those from infectious agents.
Among these vectors, adenoviral vectors, adeno-associated viral vectors and lentiviral vectors (LVs) have all been shown to give rise to transgene-directed immunity under specific conditions, in some case even being able to break pre-existing tolerance.
However, the induction of an immune response may neutralize a therapeutic protein replacing a defect protein. The unwanted immune response against antigens is also a limiting factor in conventional protein therapy (such as protein replacement therapy).
Whether the type of immune response induced by these approaches can be modulated by vector design and route of administration has long been questioned. Use of promoter targeting expression to specific cell types, such as hepatocytes, has been shown to alleviate the immune response, although it was insufficient to fully prevent its development in challenging settings.
The problem has been addressed in a number of attempts to induce immune tolerance against antigen. For example, injection into the thymus with a lentiviral vector expressing the hemagglutinin antigen (HA) in TCR-HA transgenic mice resulted in the induction of antigen-specific tolerance in the thymus and in the peripheral system (Marodon et al, 2006). Induction of immune tolerance against a specific antigen by sustained adeno-associated viral gene expression in the liver has been described for coagulation factor IX (hF.IX) (Mingozzi et al., 2003). It was also reported that expression of the neural autoantigen myelin basic protein (MBP) in the liver induced protection from autoimmune neuroinflammation in a mouse model of multiple sclerosis (Lüth et al, 2008).
We recently demonstrated that miRNA-based regulation can improve the stringency of transgene expression of LV administered to mice, is able to suppress induction of immunity against intracellular and secreted transgene products and can induce active tolerance to it. WO2007000668 describes that by using miRNA regulation to de-target transgene expression of clotting factor IX from hematopoietic lineages in hemophilia B mice it was possible to prevent immune-mediated vector clearance and enable stable gene transfer. Whereas this discovery has tremendous potential for the development of gene therapy applications as it allows for long-lasting stable expression of a therapeutic gene product delivered by a vector, the immunological outcome of tolerance induction was associated with and conceivably dependent on the stable long-term maintenance of the vector genome within the target cells, as achieved by vector integration in the case of LV. The potential dependence of the tolerogenic outcome of miRNA-regulated LV delivery from high-level vector integration and long-term robust expression within hepatocytes may significantly limit the potential applications of the discovery outside of gene replacement approaches for the correction of disease.
In particular, the efficient integration of LV within target hepatocytes may raise significant safety concerns associated with the risk of insertional mutagenesis and the triggering of oncogenesis.
Thus, the development of novel strategies that could exploit the type of antigen presentation made possible by miRNA-regulated LV without depending on or giving rise to stable long term genetic modification of recipient cells would represent a significant advancement in gene therapy.
The present invention addresses these needs and demonstrates the induction of transgene-specific active immune tolerance without the need for high level of transgene expression and vector insertion into the target cell genome.